The goals of this research are to elucidate an understanding of the molecular principles of membrane protein interactions. These interactions are important for cellular function, but they are currently poorly understood. To accomplish this goal, the stability of the transmembrane beta-barrel protein, OMPLA, will be determined. The influence of calcium and substrate, two biological effector molecules, on OMPLA self-association will also be quantified. To meet these goals, two sets of experimental studies will be carried out. Sedimentation equilibrium analytical ultracentrifugation will be used to measure OMPLA self-association equilibrium constants in the presence and absence of substrate and calcium. Spectroscopic techniques will be used to determine the strength of calcium binding to the monomeric and dimeric forms of OMPLA. To understand the mechanisms by which these three effects regulate the OMPLA activity, the thermodynamic linkage relationships between them will be determined. This thermodynamic data will be used to construct appropriate mathematical equations that describe the species populations, from which one can gain a greater understanding of the OMPLA species that are biologically relevant. The structure/energy relationships for membrane protein interactions have only been extensively studied in two helical membrane proteins. These results will be the first for a transmembrane beta-barrel protein and will facilitate a comparison of interaction motifs and principles between transmembrane alpha-helical and beta-barrel proteins.
This project will allow students to gain expertise in analytical ultracentrifugation as well as mathematics. To train young scientists, undergraduate students will be given the opportunity for a meaningful research experience. At the same time, this will have the dual benefit of offering graduate students the opportunity to learn how to mentor students and troubleshoot projects.
